K channels play an important role in cardiac cell function. The long-tern objective of this research is to understand the cellular processes by which cardiac K channels are activated and desensitized by neurotransmitters, hormones and drugs, and how these processes affect cardiac rate, force, and rhythm. In this proposal, studies will be focussed on the atrial muscarinic-gated K channel that is activated by acetylcholine (ACh) via the inhibitory GTP binding protein (Gi). Although the mechanism of activation of the muscarinic K current is known, the cellular processes underlying the subsequent rapid and slow phases of K current desensitization are completely unknown. Using whole-cell voltage- clamp and patch-clamp methods, the proposed studies will first test the hypothesis that the K current desensitization is produced by changes in the kinetics of the ACh-activated K channel opening via phosphorylation and dephosphorylation. The K current desensitization will be fully characterized in the presence and absence of agents that inhibit or activate specific protein kinases or phosphatases. Since ACh activates phospholipases, we will also examine whether certain intracellular second messengers generated via phospholipid metabolism (such as arachidonic acid or its metabolites) are involved in K current desensitization. Using agonists such as adenosine, ATP, endothelin, somatostatin and calcitonin gene-related peptide which activate the atrial muscarinic K current via separate, non-muscarinic receptors, we will investigate whether the ACh- induced K current desensitization is a homologous or a heterologous process. Finally, the role of muscarinic K current in the ACh-induced negative chronotropy and its desensitization will be studied in spontaneously contracting atrial cells by measuring beating rates under various conditions described above that modifies the K channel function. These studies will provide important new knowledge on the cellular mechanisms of rapid and slow desensitization of the muscarinic K current. The studies will understand the role of phosphorylation and dephosphorylation by protein kinases and phosphatases and the role of second messengers generated via phospholipases in K channel desensitization. The role of muscarinic K current in ACh-induced changes in contractile behavior will be identified.